Anti-inflammatory effects of topical supernatant from human amniotic membrane cell culture on canine deep corneal ulcer after human amniotic membrane transplantation

The objective of this study was to examine the effect of topically applied human amniotic epithelial cell (HAEC) culture supernatant on corneal inflammatory reaction in dogs. Twenty-five dogs were randomly assigned into five groups. The control group consisted of five dogs with normal cornea. Inductions of corneal ulcers were performed using 0.45 cm trephine and human amniotic membrane was transplanted in 20 dogs. These 20 dogs were assigned into four treatment groups: topical antibiotic, topical corticosteroid, topical mock media and topical culture supernatant from HAEC, respectively. Administrations of the testing agents started at 24 h (h) after transplantation four times daily for nine consecutive days. Tears were collected before an operation 24 h after transplantation, but before application of the testing agents on consecutive odd days following transplantation. The concentrations of interleukin-1 beta (IL-1 beta) and nitric oxide (NO) in tear fluid were measured using canine IL-1 beta ELISA kit and Griess assay, respectively. Our analysis indicates that elevations of IL-1 beta and NO concentrations are associated with inflammatory conditions in the eyes. Corticosteroid, a reference anti-inflammatory drug, and the culture supernatant from HAEC significantly decreased IL-1 beta and NO concentrations. In addition, the clinical signs such as conjunctivitis and neovascularization were decreased in both topical corticosteroid and supernatant from HAEC treated groups. Mock and antibiotic solutions failed to decrease NO and IL-1 beta concentrations. In conclusion, topical application of the culture supernatant from HAEC alleviated inflammation in induced-corneal ulcer of dogs, possibly via inhibition of IL-1 beta and NO production.